Contrast agents are used to assist in the identification of molecular and supramolecular targets or regions with distinctive local state properties. Non-limiting examples of targeted species are proteins, polysaccharides, polynucleic acids and other analytes, or environmental properties. Non-limiting examples of state properties include distinct pH, temperature, and solvent quality. The targeted molecule or state may be difficult to observe in the background of other material, and the contrast agent makes it easier to identify it. One example is a particular type of cell surface protein in the presence of other types of cell surface proteins. Often such proteins are the markers of a health condition, and their identification can assist in diagnosis, determination of treatment or monitoring the progress of a health condition. Contrast agents can be used to detect other species, such as chemical threats or biological and chemical warfare agents.
Optical contrast agents, which function using light, are attractive because the technology to employ them can be relatively simple, for example a light microscope or spectrometer coupled to a sampling device. Brighter optical contrast agents can be easier to detect than less bright agents. Methods to create brighter particles are therefore valuable. It is useful to have contrast agents that can be easily multiplexed, meaning that multiple kinds of contrast agents can be used on the same sample to detect simultaneously the presence and number of multiple targets. For example, the type and number of more than one type of protein might be detected on the surface of a single cell, which could improve in identifying the state of the cell and its relevance to the health of the individual. Optical contrast agents that can be multiplexed, which means optical contrast agents that can be used in the presence of other different contrast agents, are therefore valuable.
Plasmonic nanomaterials, and their utility in surface enhanced chemical and biological sensors, have garnered immense interest in the past two decades due to their size-dependent optical properties and potential as target specific contrast agents. Surface-enhanced Raman spectroscopy (SERS) is the most widely studied and offers the possibility of single molecule detection.
Raman scattering nanoparticles are potentially useful as optical contrast agents because they exhibit sharper optical signatures than, for example, most fluorescent contrast agents. This means that they can in principle be highly multiplexed. Raman scattering by isolated molecules is, however, weak, and therefore methods to create brighter or more strongly scattering Raman contrast agents would be useful.
Raman-scattering nanoparticles have great promise as sensitive detection labels. However, due to complex design criteria such as binding specificity, robust colloidal stability in biological environments, and optical sensitivity, few commercially viable sensor systems have been generated as a result of this widespread research.
Active plasmonics, defined as plasmonic structures coupled to materials that can interact with the plasmon, has generated significant interest, especially in the past few years, as a next generation platform to address the need for brighter SERS signals. J-aggregates, Rhodamine 6G, cytochrome c, porphyrin derivatives, and host-guest charge transfer complexes have all been successfully coupled to plasmonic nanostructures to elicit a much brighter surface-enhanced resonance Raman (SERRS) signal. Even greater enhancement has been achieved when a wavelength matching approach is employed to couple the molecular and plasmonic resonances together with the excitation field. Appropriate coordination of multiple resonating entities offers a powerful tool for brighter SERS-based sensing platforms. However, while these other approaches have demonstrated some improvement to the optical brightness, incorporation into a robust and stable detection label modality has not been accomplished.
For practical use of nanoparticles as a detection modality, stability in various solutions and shelf life are of paramount importance. While in water, these particles may exhibit desirable properties, but when placed in biological media or buffer solution, they tend to aggregate very quickly and optical properties are greatly diminished as a result. Often, smaller particles are employed during complicated surface chemistry, as they are more resistant to aggregation due to electrostatics, i.e. when charged species or proteins are introduced into solution and around the particle. However, smaller particles have much weaker plasmon resonances, so there is an effective trade-off between particle stability and effective optical brightness to make the composite particles useful as contrast agents.
Several ways to increase the long term stability of larger SERS-active nanoparticles have been proposed, the most popular being the co-adsorption of various PEG chain lengths to the surface of the particle along with the Raman dye. Recently, encapsulation of the Raman dye within a stabilizing to coating layer that surrounds particles has been employed for relatively large particles of 60 nm in diameter. These stabilizing layers can consist of inorganic oxides such as silicon or titanium, or organic self-assembled structures such as lipid vesicles. These composite particles have shown great promise as shelf-stable, biologically compatible biosensors, as the stabilizing coating prevents particle aggregation in biological media while offering a versatile platform for targeting functionality and other surface chemistry.